| The term’endophytes’refers to any organisms (bacteria or fungi) occurring within plant tissues without causing apparent hurt to that plant or showing any signs of existence in that plant. Endophytic fungi are ubiquitous and have been found in all plant species examined to date. Endophytic fungi are hyperdiverse, also. Simultaneously, endophytic fungi have diverse functional roles. In this study, the endophytic fungi were isolated from oilseed rape (Brassica napus) and identified based on morphological chacterieristics and rDNA-ITS. Biocontrol efficacy of these fungal isolates was evaluated in vitro and in vivo. The main results were summarized below:1. The endophytic fungi were isolated from root, stem and leaf of healthy oilseed rape. The endophytic fungi were identified based on morphology together with ITS sequences. Distribution of endophytic fungi in different organs (root, stem, leaf) and at different growing stage (seedling, budding, flowering and pod-development) was analyzed. The Shannon index (H’) and Simpson’s diversity index (1-D) were calculated. Total of97endophytic fungal isolates were obtained from roots (35), stems (49) and leaves (13) of oilseed rape. Forty fungal species were identified and most species (80%) belong to Ascomycota. The Simpson’s diversity index of root, stem and leaf were0.936,0.962and0.731, respectively.The all species composition is highly diversified with Simpson’s diversity index reaching0.959. The result indicated the diversity of ednophytic fungi of oilseed rape. The specieses Alternaria alternate and Chaetomium globosum existed in root, stem and leaf. Five specieses Macrophomina sp., Aspergillus versicolor, Penicillium pinophilum, Fusarium tricinctum and Fusarium oxysporum exist in roots only.2. Twenty-four isolates exhibited antifungal activity against S. sclerotiorum in dual cultures on potato dextrose agar plate forming inhibition zones of3-17mm in width. These strains included Alternaria, Aspergillus, Chaetomium, Clonostachys, Dothidea, Epicoccum, Fusarium, Leptosphaeria, Simplicillium genera. The culture filtrates of Aspergillus flavipes CanS-34A, Chaetomium globosum CanS-73, Clonostachys rosea CanS-43and Leptosphaeria biglobosa CanS-51in potato dextrose broth exhibited consistent and effective suppression of oilseed rape leaf blight caused by S. sclerotiorum. In addition, six strains were found to be able to inhibit S. sclerotiorum growth by producing volatile organic compounds. 3. Thirty-five endophytic strains were found that could produce IAA. These fungi included Alternaria, Aspergillus, Aureobasidium, Chaetomium, Epicoccum, Fusarium, Hypoxylon, Macrophomina, Nigrospora, Periconia, Phoma and Sporidiobolus sp. The cultural filtrates of the strains Fusarium proliferatum CanR-9, Sporidiobolus sp. CanS-62and Dioszegia zsoltii CanS-69were found capable of promoting lateral root growth of oilseed rape. A. alternata CanL-18, Fusarium tricinctum CanR-70and CanR-71r, and L. biglobosa CanS-51exhibited growth-promoting effects on oilseed rape during the two-year pot experiment.4. The antifungal substances of Aspergillus flavipes CanS-34A have been studied. The cultural filtrates of A. flavipes CanS-34A inhibited15plant pathogenic fungi. The percentage of growth inhibition was higher for four Sclerotinia species (S. sclerotiorum, S. trifoliorum, S. minor and S. nivalis)(74.9%-100%) than for the other11fungi (13.9%-71.9%). The antifungal activity of cultural filtrates was found resistant to heat treatment below100℃.5. Nineteen compounds were identified in the VOCs emitted by non-pathogenic Fusarium oxysporum CanR-46via GC-MS. The Fo-VOCs inhibited growth of all the16tested fungal species, delayed conidial germination, and suppressed germ-tube elongation of Be and Vd. Damaged (shriveled and collapsed) mycelia of both Be and Vd caused by the Fo-VOCs were observed under scanning electron microscope. Four (5-hexenoic acid, limonene,3,4-2H-dihydropyran and octanoic acid) synthetic chemicals present in the Fo-VOC profile showed antifungal activity against Be and Vd. The VOCs emitted from Fo cultures significantly (P﹤0.05) reduced disease severity of Botrytis fruit rot of tomato compared to the treatment Be alone. Treatment of cotton roots with Fo provided effective control of Verticillium wilt of cotton compared to the treatment with Vd alone. Colonization of the epidermis and vascular tissues of cotton roots by a GFP-transformed strain of Fo was consistently observed under confocal laser scanning microscope.These results might be useful for utilization of the endophytic fungi to control diseases of oilseed rape in the future. |
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